Cells adhere to extracellular matrix (hereinafter abbreviated as ECM) mediated by a group of cell surface receptors which are termed integrins. Integrins perform their functions by forming 1:1 heterodimers of α and β chains. At least 18 types of α chain, 8 types of β chain and 24 types of αβ heterodimer have been identified and confirmed so far. It is known that each integrin recognizes a specific ligand. Integrins are classified into subfamilies depending upon the specificities or functions to ligands, and divided into collagen receptors, laminin receptors, RGD receptors recognizing an Arg-Gly-Asp (RGD) sequence present in fibronectin, vitronectin, etc., leukocyte-specific receptors present only in leukocytes (Non-Patent Literature 1: Hynes, R. O., 2002, Integrins: Bidirectional, Allosteric Signaling Machines. Cell 110: 673-87; Non-Patent Literature 2: Miyasaka, M., 2000, New edition of Adhesion Molecule Handbook, Shujunsya). The α4 and α9 integrins are a subfamily that does not belong to any of these types and called the α4 integrin subfamily (Non-Patent Literature 3: Elise L. Palmer, Curzio Rfiegg, Ronald Ferrando, Robert Pytela, Sheppard D., 1993, Sequence and Tissue Distribution of the Integrin α9 Subunit, a Novel Partner of β1 That Is Widely Distributed in Epithelia and Muscle. The Journal of Cell Biology, 123: 1289-97). On the other hand, ECM was considered so far to serve as a mere cementing substance between cells. It has now become clear that the integrin-mediated ECM-cell interaction is deeply involved in regulating the growth, adhesion, movement, etc. of cells and associated with the onset of diseases including a progression of cancer, an exacerbation of inflammation, etc.
Osteopontin (hereinafter abbreviated as OPN) which is one of ECM is a secreted, acidic phosphorylated glycoprotein with a molecular weight of about 41 kDa and is a molecule, which expression is widely observed in breast milk, urine, renal tubules, osteoclasts, osteoblasts, macrophages, activated T cells, tumor tissues, etc. OPN has the adhesion sequence GRGDS (SEQ ID NO: 16) at the center of its molecule, the SVVYGLR sequence (SEQ ID NO: 15) in human OPN or the SLAYGLR sequence (SEQ ID NO: 18) in mouse OPN and a thrombin-cleavage site in close proximity thereto, and binds through the GRGDS sequence (SEQ ID NO: 16) to the RGD integrin or to the α4 (α4β1) and α9 (α9β1) integrins through the SVVYGLR sequence (SEQ ID NO: 15) or the SLAYGLR sequence (SEQ ID NO: 18).
Differences in binding profile are also found in that α4β1 binds both to OPN not cleaved with thrombin (uncleaved OPN) and to the N-terminal fragment of thrombin-cleaved OPN (cleaved OPN), whereas α9β1 binds only to the cleaved OPN (Non-Patent Literature 4: Y. Yokosaki, et al., (1999) The Journal of Biological Chemistry, 274: 36328-36334; Non-Patent Literature 5: P. M. Green, et al., (2001) FEBS Letters, 503: 75-79; Non-Patent Literature 6: S. T. Barry, et al., (2000) Experimental Cell Research, 258: 342-351).
The α4 and α9 integrins share many common ligands other than OPN. Known ligands are the EDA domain of fibronectin, propeptide-von Willebrand factor (pp-vWF), tissue transglutaminase (tTG), blood coagulation factor XIII, vascular cell adhesion molecule-1(VCAM-1), etc. In addition, the CS-1 domain of fibronectin, MadCAM-1 (α4β7), etc. are known as the ligands specifically recognized by the α4 integrin. Tenascin-C, plasmin, etc. are known as the ligands specifically recognized by the α9 integrin.
The amino acid sequences for the integrin subunits α9, α4 and β1 are publicly known. For instance, human α9 is registered as NM—002207, mouse α9 as NM—133721, human α4 as NM—000885, mouse α4 as NM—010576, human β1 X07979, and mouse β1 as NM—010578, at the GenBank. These integrins are also known to have high similarities between species in amino acid sequence.
WO 02/081522 (Patent Literature 1) discloses a therapeutic effect on rheumatoid arthritis or hepatitis by inhibiting the OPN functions using OPN knockout mice or neutralizing antibodies against OPN. Moreover, this patent literature discloses that the SVVYGLR sequence (SEQ ID NO: 15) is essential as recognizing the α9 and α4 integrins for pathogenesis of an inflammatory disease and that receptors for OPN are expressed in immunocompetent cells or the like and associated with an inflammatory disease.